Microorganisms from the genus Salmonella are responsible for the majority of cases of bacterial diarrhea occurring in humans. Typically, screening and identification of a Salmonella infection is accomplished by microbial culturing techniques followed by biochemical testing. This takes from one to three days, sometimes ending in equivocal results. Identification of Salmonella spp. by use of an assay using DNA probes would be less labor intensive for the clinician, provide unequivocal results, and would enable quicker diagnosis of salmonellosis.
Nucleic acid hybridization technology is a new approach in the diagnostic industry. This methodology exploits the property that sequences unique to an organism comprise its genome. In a hybridization test, a positive signal is generated when these unique genomic sequences in the bacterial or the viral target hybridize with nucleic acid probes, which are tagged with a detection label. U.S. Pat. No. 4,689,295 to Taber, et al. describes Salmonella DNA probes which are Salmonella DNA fragments common to more than 80% of the known Salmonella species. The fragments do not code for any protein or any other material known to contribute to pathogenicity. Thus, the probes are constructed from a library of fragments without regard to genetic function, and are principally used to detect the presence of Salmonella in a food sample.
U.S. Pat. No. 4,358,535 to Falkow, et al. describes a method for detecting microorganisms using a DNA probe by affixing the genetic material from a sample containing the microorganism to an inert support and treating the sample to render the genetic material in a single-stranded form. The fixed material is then contacted with a labeled nucleotide sequence that is substantially complementary to a nucleotide sequence of a structural gene encoding for toxins produced by the microorganisms. E. coli is the only organism exemplified in the patent, although Salmonella is mentioned in a laundry list of toxin-producing microorganisms.
U.S. Pat. Nos. 4,486,539 and 4,563,419 to Ranki, et al. describe a one-step sandwich hybridization technique employing reagents having distinct single-stranded nucleic acid fragments isolated from the genome of a microbe to be identified. These fragments have no extensive sequence homology but are situated closely together in the genome. There is no mention of specific genes encoding for a proteinaceous virulence factor that may be involved in human pathogenicity from infection with a microbe from the genus Salmonella.
Nichols and Yanofsky, Proc. Natl. Acad. Sci., Vol. 76, No. 10, pp. 5244-5248 (October 1979) describe the nucleotide sequences of trpA of Salmonella typhimurium and Escherichia coli and make an evolutionary comparison of the two. There is no mention of using these sequences as DNA probes.
Stoleru, et al., Chem. Abstracts, 85:90053b (1976) describe polynucleotide divergence among strains of Salmonella sub-genus IV and other closely related organisms. Here again, there is no mention of using sequences for DNA probes.